Neuronal development and regeneration are complex processes guided by neurotrophic factors, including neuropeptides. The vasoactive intestinal peptide (VIP/pituitary adenylyl cyclase activating polypeptide (PACAP) family of neuropeptides is implicated in many aspects of development. The overall goal of this application is to study the roles of PACAP and its receptors in neuronal proliferation and maturation using olfactory receptor neurons (ORNs) as a model. While their localization during embryogenesis suggests a critical role for PACAP peptides, the mechanisms whereby PACAP mediate these processes is not understood. The olfactory system has several attributes that make it attractive for modeling neuropeptide function. In addition to a developmental program, ORNs regenerate post-natally from maintained precursor cells. Our data demonstrate that ORNs express high levels of PACAP and its receptors during development, making them likely mediators of developmental signals. In addition, PACAP affects ORN proliferation in vitro. The use of partial loss of function knockout models confirms this role in vivo. We hypothesize that PACAP is essential to ORN development and transduces proliferation or differentiation signals through the coupling of PACAP receptors to specific intracellular cascades. Aim I will characterize PACAP and its receptor expression profiles developmentally, during adulthood, during regeneration post-lesioning of the ORN target (the olfactory bulb), and in primary ORN cultures. We will determine which cells types express PACAP and its receptors in different developmental and post-lesioning paradigms and which cell types in the olfactory epithelium express PACAP peptides. Aim II will investigate the roles of PACAP in ORN proliferation, survival and differentiation. We will use PACAP and its antagonist, BrdU/PCNA labeling, TUNEL assays, primary ORN cultures and ORN cell lines as well as adenovirus-mediated gene transfer of antisense RNA to study the roles of PACAP and its receptors in O4RN proliferation and survival in vitro. In Aim Ill, we will use pharmacological reagents and biochemical second messenger assays in ORN primary cultures and cell lines to study the intracellular signaling cascades activated by PACAP during different stages of ORN development. We will determine if PACAP peptides modulate ORN proliferation and differentiation by regulating cyclin-dependent kinases and the inhibitory protein CIP/KIP. These studies will elucidate the function and mechanisms underlying bioactive peptide neurotrophism in the CNS.